Mycotoxins are noxious compounds produced by a variety of fungal species. One such mycotoxin, aflatoxin, has been linked to contaminated food (e.g., corn, rice, peanuts, and tree nuts) and animal feed (Woloshuck 1998, Smela 2001). Aflatoxins are produced by specific strains of the filamentous fungus Aspergillus (aflatoxin=Aspergillus flavus toxin) and encompass a group of structurally related compounds (Trail 1995). One member in particular, aflatoxin B1 (AFB1), is the most toxic and the most prevalent in nature (Woloshuck 1998). Up to twenty five percent of the global food supply is contaminated, annually, by AFB1 (Trail 1995, Moreno 1999). In the United States, the Food and Drug Administration (FDA) regulates the levels of aflatoxin in foods such that crops with more than 20 parts per billion (ppb) total aflatoxins cannot be imported/exported and sold (Gourama 1995, Trail 1995).
Aflatoxins induce specific point mutations in DNA. Typically, aflatoxins like AFB1 are metabolized in the liver. There they are converted into epoxides (AFB-8,9-epoxide) which subsequently become covalently linked to guanine bases in the liver cell DNA (Eaton 1994, Wang 2000, Smela 2001). Addition of the epoxide (usually at the N7 position) stimulates depurination of the guanine base which is then misinterpreted during subsequent DNA replication (Smela 2001). Thus, aflatoxin induces GC→TA transversions within the DNA. One such point mutation has been shown to readily form within the liver p53 tumor suppressor gene (G249T) and in fact, this particular transversion has been directly correlated with the occurrence of hepatocellular carcinoma (i.e., liver cancer) (Hussain 1994, Moreno 1999, Tiemersma 2001).
In recent years, strategies have been proposed to eliminate aflatoxins from food and feed. In the field, application of fungicide has prevented fungal infection and subsequently, mycotoxin contamination. Aflatoxin detection via chromatography and UV luminescence coupled with post-harvest removal techniques have also been utilized (Kathuria 1993). These current elimination strategies, however, have been costly, ineffective, and/or environmentally unsound (Trail 1995, Campbell 2003). Thus, there is a need for simpler, less expensive ways of limiting or preventing aflatoxin contamination of food and feed.
Alfalfa (Medicago sativa) is the most widely grown forage in the western United States, providing over $2 billion of cash income to its producers. Alfalfa constitutes about 23 to 34% of dairy ration on dry matter basis (Getachew et al. 2005). However, alfalfa protein and nitrogen (N) utilization by ruminants is generally considered to be sub-optimum. Over 50% of the N contained in alfalfa forage is utilized poorly, or not at all, for animal products (e.g., milk, meat, fiber). The result is excretion of excess N, one of the most significant non-source groundwater and air contamination sources. Because of poor N utilization, high producing dairy cows require costly protein concentrate supplements. Protein concentrate production itself is water and energy intensive. Improving protein utilization and feeding value of alfalfa must be considered as the single highest impact intervention for environmental compatibility and sustainability of dairy production. Excess N from high producing dairy cows is one of the most critical environmental problems—not only in California, but in all dairy regions of the world. There is, thus, a need for plants with improved protein utilization characteristics.